1. Field of the Invention
The present invention relates to simultaneous optical detection of two or more parameters of dissolved oxygen concentration, pH and temperature and, more particularly, to an optical sensing membrane, comprising a mixture containing two or more fluorescent dyes for detection of dissolved oxygen concentration, pH and temperature, immobilized on a support, a detection device including the optical sensing membrane, and a detection method using the detection device.
2. Description of Related Art
Dissolved oxygen concentration, temperature and pH are important parameters in fields of environmental monitoring, ocean research, food industries, bioengineering, and medicines. Optical detection has an advantage over other methods since it can be carried out in a non-invasive manner, through a bioreactor or a reaction vessel of glass. In this case, sensing materials are positioned on an inner wall of the reaction vessel and detection is carried out by reflection or fluorescence in the outside of the reaction vessel. Further, the optical sensor has another advantage not to have electromagnetic fields generated since it measures signals using the light instead of electronic element and transfers the measured information using the light. In particular, it is widely used in various fields of sensors due to development of optical dyes selectively emitting the light depending upon specific materials, e.g., dissolved oxygen molecules, carbon dioxide molecules, and the like, and pH variance. Such optical analysis is widely used in various fields since it meets economic factors such as time and cost reduction. Recently, a compact multiple bioreactor has been actively developed using a compact reactor and an optical monitoring technique. The compact multi-bioreactor has an advantage of minimizing costs for process development, since it enables the development of desired process conditions at low costs for a short period of time in the optimization of production process of pharmaceuticals and biotechnological products.
Fluorescent dyes capable of sensing dissolved oxygen concentration, pH and temperature include Rudpp (tris-4,7-dipenyl-1,10-phenanthroline) ruthenium (II) complex, HPTS (8-hydroxypyrene-1,3,6-trisulfonic acid trisodium salt), Quantum dots (Qds) and rhodamine B. Rudpp has a property of emitting fluorescence of 600 nm when an excitation light of 480 nm is incident, in which the fluorescence is generated in inverse proportional to dissolved oxygen concentration. HPTS has a property of emitting fluorescence of 520 nm when an excitation light of 410 nm is incident, in which the fluorescence intensity is increased as a hydrogen ion concentration is decreased. Quantum dots and rhodamine B are used as fluorescent dyes for measuring temperature.
Simultaneous detection of two or more of dissolved oxygen concentration, pH and temperature is required in a complicated cultivation and process. Further, since a quenching effect of dissolved oxygen concentration is greatly influenced by temperature, it is important to know temperature in the optical detection of dissolved oxygen concentration. Recently, fluorescent dyes for simultaneous optical detection of two parameters have been reported. For example, “Talanta 47 (1998) pp. 1071-1076” describes a technique of immobilizing ferrozine, an iron indicator, and HPTS, a pH indicator, in adjacent areas of a single thin membrane, to detect iron and pH in two distinguishable wavelengths by attenuated total reflection (ATR) spectroscopy. “Chem. Mater. 2006 pp. 4609-4616” describes a single-fiber optical sensor capable of simultaneously detecting pH and dissolved oxygen concentration, which has carboxyfluorescein as a pH probe and Rudpp as a dissolved oxygen probe, respectively. However, they have never disclosed the combination of two or more fluorescent dyes of Rudpp, HPTS, and quantum dots or rhodamine B. Further, “Analytical Chimica Acta 606 (2008) pp. 223-229” discloses a technique of detecting temperature and dissolved oxygen concentration using quantum dots and Rudpp, but it concerns the immobilization of quantum dots and Rudpp in adjacent areas on a single thin membrane (See FIGS. 1 and 2.1. Materials therein), which is completely different from making a homogeneous mixture and following immobilization of the mixture.